PROJECT SUMMARY - OVERALL We propose preclinical and clinical studies of oncolytic Herpes simplex viruses (oHSVs) for the therapy of glioblastoma (GBM), a deadly and incurable neoplasm. This class of biologic agents (recently approved by the FDA for melanoma) are thought to exert their effects through two mechanisms: there is an initial phase of OV infection of tumor cells, followed by serial rounds of viral progeny production and spread of the infection to adjacent tumor cells. This first phase results in direct tumor cytotoxicity, produces inflammatory responses and release of tumor antigens that activate a second phase of cytotoxic T cell responses, responsible for the durable anticancer activity of virotherapy. During the current cycle, we discovered that the initial phase of oHSV action is hindered by rapid and premature clearance of oHSVs by cells of the innate immune system, i.e., natural killer (NK) cells and macrophages. Our overall hypothesis is that NK cells and macrophages are called into the GBM microenvironment to rapidly remove oHSV-infected tumor cells, before sufficient viral replication and tumor cytotoxicity occurs. Recognizing that this hypothesis requires broadening based on the clinical success of immune checkpoint (IC) blockade, a corollary is that oHSV-mediated immunostimulation and GBM inflammation when coupled with restoration of T cell activation (via immune checkpoint blockade) leads to a durable ?anti-GBM? effect. Our four projects propose two overall approaches to test these hypotheses: (a) direct inhibition of NK cell receptor activation against viral antigens on tumor cell surfaces (Project 4-Dr. Caligiuri; The Ohio State University Comprehensive Cancer Center, OSUCCC) and/or inhibition of oHSV-infected tumor cell signaling that activate NK cells (Project 1-Dr. Glorioso; University of Pittsburgh Medical Center, UPMC) and macrophages (Project 3- Dr. Kaur, University of Texas Health Sciences, UTHealth), and (b) addition of immune checkpoint inhibition to oHSV therapy (Project 2-Dr. Chiocca, Brigham and Women's Hospital/Dana Farber Cancer Institute, BWH/DFCI). Further, this PPG will perform a ?first-in-man? clinical trial in recurrent GBM patients using an oHSV whose development was completed during the current funding period. Project 2 will provide the clinical samples to the other Projects and Cores to allow for the clinical validation of experimental outcomes. Three Cores provide oHSV preparation (Core 1- Dr. Goins, UPMC), mouse and human GBM cells (Core 2-Dr. Ligon, DFCI) and biostatistical analyses (Core 3-Dr. Fernandez, OSUCCC). Our group comprises experts in virology, cancer biology, immunology, and clinical trials with biologic therapies and is uniquely positioned to assess the preclinical and clinical efficacy of oHSV therapy for patients with recurrent GBM.